Crystal Structures Reveal the Multi-Ligand Binding Mechanism of Staphylococcus aureus ClfB

Staphylococcus aureus (S. aureus) pathogenesis is a complex process involving a diverse array of extracellular and cell wall components. ClfB, an MSCRAMM (Microbial Surface Components Recognizing Adhesive Matrix Molecules) family surface protein, described as a fibrinogen-binding clumping factor, is a key determinant of S. aureus nasal colonization, but the molecular basis for ClfB-ligand recognition remains unknown. In this study, we solved the crystal structures of apo-ClfB and its complexes with fibrinogen α (Fg α) and cytokeratin 10 (CK10) peptides. Structural comparison revealed a conserved glycine-serine-rich (GSR) ClfB binding motif (GSSGXGXXG) within the ligands, which was also found in other human proteins such as Engrailed protein, TCF20 and Dermokine proteins. Interaction between Dermokine and ClfB was confirmed by subsequent binding assays. The crystal structure of ClfB complexed with a 15-residue peptide derived from Dermokine revealed the same peptide binding mode of ClfB as identified in the crystal structures of ClfB-Fg α and ClfB-CK10. The results presented here highlight the multi-ligand binding property of ClfB, which is very distinct from other characterized MSCRAMMs to-date. The adherence of multiple peptides carrying the GSR motif into the same pocket in ClfB is reminiscent of MHC molecules. Our results provide a template for the identification of other molecules targeted by S. aureus during its colonization and infection. We propose that other MSCRAMMs like ClfA and SdrG also possess multi-ligand binding properties

Functional Analysis of the Arlequin Mutant Corroborates the Essential Role of the ARLEQUIN/TAGL1 Gene during Reproductive Development of Tomato

Reproductive development of higher plants comprises successive events of organ differentiation and growth which finally lead to the formation of a mature fruit. However, most of the genetic and molecular mechanisms which coordinate such developmental events are yet to be identified and characterized. Arlequin (Alq), a semi-dominant T-DNA tomato mutant showed developmental changes affecting flower and fruit ripening. Sepals were converted into fleshy organs which ripened as normal fruit organs and fruits displayed altered ripening features. Molecular characterization of the tagged gene demonstrated that it corresponded to the previously reported TOMATO AGAMOUS-LIKE 1 (TAGL1) gene, the tomato ortholog of SHATTERPROOF MADS-box genes of Arabidopsis thaliana, and that the Alq mutation promoted a gain-of-function phenotype caused by the ectopic expression of TAGL1. Ectopic overexpression of TAGL1 resulted in homeotic alterations affecting floral organ identity that were similar to but stronger than those observed in Alq mutant plants. Interestingly, TAGL1 RNAi plants yielded tomato fruits which were unable to ripen. They displayed a yellow-orange color and stiffness appearance which are in accordance with reduced lycopene and ethylene levels, respectively. Moreover, pericarp cells of TAGL1 RNAi fruits showed altered cellular and structural properties which correlated to both decreased expression of genes regulating cell division and lignin biosynthesis. Over-expression of TAGL1 is able to rescue the non-ripening phenotype of rin and nor mutants, which is mediated by the transcriptional activation of several ripening genes. Our results demonstrated that TAGL1 participates in the genetic control of flower and fruit development of tomato plants. Furthermore, gene silencing and over-expression experiments demonstrated that the fruit ripening process requires the regulatory activity of TAGL1. Therefore, TAGL1 could act as a linking factor connecting successive stages of reproductive development, from flower development to fruit maturation, allowing this complex process to be carried out successfully